Oligocone Trichromacy: Clinical and Molecular Genetic Investigations

Clinical and Epidemiologic Research Oligocone Trichromacy: Clinical and Molecular Genetic Investigations

Mette K. G. Andersen,1,2 Nynne L. B. Christoffersen,1 Birgit Sander,3 Carsten Edmund,4 Michael Larsen,1,3 Tanja Grau,5 Bernd Wissinger,5 Susanne Kohl,5 and Thomas Rosenberg1,2

PURPOSE. To describe the phenotype and genotype of patients ligocone trichromacy (OT) is a rare form of congenital with a diagnosis of oligocone trichromacy (OT). Ocone dysfunction characterized by a paradoxical combi- METHODS. Six unrelated patients had a detailed ophthalmic nation of severe electrophysiological cone impairment and normal or near-normal color discrimination. OT was delineated examination including color vision testing, a Goldmann visual 1 field test, fundus photography, and full-field electroretinogra- as a nosologic entity by van Lith, who reported on a patient phy (ffERG). Five patients also underwent multifocal (mf)ERG, with congenital nystagmus and low vision. The condition was autofluorescence recording, and optical coherence tomogra- further characterized by a severely reduced photopic electro- phy (OCT). Genetic analysis included sequencing of all coding retinogram (ERG) response and the startling finding of almost regions and flanking introns of CNGA3, CNGB3, GNAT2, normal color vision. Further examinations revealed normal KCNV2, and PDE6C. peripheral visual fields, elevated cone thresholds, lowered flicker fusion frequency, normal or moderately reduced rod RESULTS. All patients had subnormal visual acuity, a history of ERG, and normal electrooculogram (EOG). Based on these congenital nystagmus, and subjectively normal or near-normal observations van Lith hypothesized a very low number of color vision; five patients reported photophobia. Clinical ex- normal functioning cones, explaining both the low ERG cone aminations revealed largely normal fundi, normal Goldmann response and the near normal color vision. Keunen et al.2 visual field results with the IV/4e target, and normal color corroborated this hypothesis by measuring foveal cone pho- discrimination or mild color vision deficiency. Electrophysio- topigment density in four individuals with OT and found a logical investigations showed either complete absence of re- reduced density difference but a normal photopigment regen- cordable cone responses or severely reduced amplitudes. All eration time constant. retinal layers were identifiable by OCT, which also showed Taxonomically OT is classified as a cone dysfunction, a very thinning of the peripheral retina. Genetic analysis revealed two heterogeneous group of disorders including total and incom- causative CNGB3 mutations in one patient and single heterozy- plete achromatopsia (rod monochromatism), blue cone mono- gous mutations of unknown significance in CNGB3 and PDE6C chromatism, Bornholm eye disease, and cone monochroma- in two other patients. tism. (For a recent review, see Michaelides et al.3 and CONCLUSIONS. Oligocone trichromacy is a heterogeneous con- references therein.) dition with respect to both phenotypic appearance and genetic In this report, we present multimodal clinical findings and background. The finding of mutations in genes known to be the results of molecular genetic investigations in six patients involved in complete and incomplete achromatopsia supports with a clinical diagnosis of OT. the notion that some forms of OT is an extreme form of incomplete achromatopsia with preferential loss of peripheral cones. (Invest Ophthalmol Vis Sci. 2010;51:89–95) DOI: METHODS 10.1167/iovs.09-3988 All patients receiving a diagnosis of OT at the National Eye Clinic from 1993 to 2001 were invited to participate in the study. Inclusion criteria were a history of congenital nystagmus, subnormal visual acuity (0.9– From the 1National Eye Clinic and 2The Gordon Norrie Centre for 0.1), subjectively normal or near-normal color vision, and absent or 3 Genetic Eye Diseases, Kennedy Center, Glostrup, Denmark; the De- severely reduced ERG cone responses. Six unrelated patients volun- partment of Ophthalmology, Glostrup Hospital, University of Copen- 4 teered and gave their informed consent in compliance with the tenets hagen, Glostrup, Denmark; the Department of Ophthalmology, Rig- of The Declaration of Helsinki. Most of the patients reported having shospitalet, University of Copenhagen, Copenhagen, Denmark; and the photophobia since childhood. Five of the six had been referred as 5Molecular Genetics Laboratory, Institute for Ophthalmic Research, Centre for Ophthalmology, University Tu¨bingen, Tu¨bingen, Germany. children with congenital nystagmus and reduced visual acuity. Patient Supported by a scholarship from the Danish Agency for Science B, with near-normal visual acuity, and minimal nystagmus was referred Technology and Innovation (MKGA). The molecular genetic analysis at age 40. was supported by German Research Council (DFG) Grant KFO134- The study protocol included refraction and assessment of best- Ko2176/1. corrected visual acuity, Goldman manual kinetic perimetry (I/4e and Submitted for publication May 14, 2009; revised June 22 and July IV/4e targets), slit lamp biomicroscopy with fundus examination, eval- 1, 2009; accepted July 5, 2009. uation of nystagmus, ERG, color vision testing, optical coherence Disclosure: M.K.G. Andersen, None; N.L.B. Christoffersen, tomography (OCT), fundus autofluorescence imaging, and color fun- None; B. Sander, None; C. Edmund, None; M. Larsen, None; T. dus photography. Patient F was unavailable for some of the proce- Grau, None; B. Wissinger, None; S. Kohl, None; T. Rosenberg, None dures. Corresponding author: Thomas Rosenberg, Gordon Norrie Centre Color vision testing comprised Farnsworth-Munsell’s 100-hue test for Genetic Eye Diseases, Gl. Landevej 7, DK-2600 Glostrup, Denmark; (FM-100), Farnsworth-Munsell’s D-15 (FM D-15, saturated), Lanthony’s [email protected]. tritan album (LTA), and Ishihara’s 38 pseudoisochromatic plates (2001

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TABLE 1. Summary of Clinical Findings

Sex, Age at Time of Follow-up BCVA Refraction Patient Follow-up Period (y) (Right/left) (Right/left) Nystagmus Photophobia Goldmann Perimetry

A M, 46 32 0.4/0.5 ϩ1.00/ϩ0.50 ϩϩ No Normal boundaries for target IV/4e and I/4e, no scotoma B F, 47 8 0.6/0.9 ϩ3.00/ϩ2.75 ϩ Yes Normal boundaries for target I/V4e, no scotoma, cannot see target I/4e C M, 28 26 0.2/0.2 0/0 ϩϩϩ Yes Normal boundaries for target IV/4e and I/4e, no scotoma D F, 12 12 0.1/0.2 Ϫ2.00/Ϫ4.25 ϩ Yes Normal boundaries for target IV/4e, no scotoma; constricted ﬁeld with I/4e EF,248Ͻ0.6/0.5 ϩ1.25/ϩ0.75 None (history of Yes Normal boundaries for target congenital IV/4e and I/4e, no nystagmus) scotoma F M, 8 None 0.3/0.3 ϩ1.50/ϩ1.00 ϩϩ Yes Normal boundaries for target IV/4e, no scotoma

edition). Two patients were also examined with Nagel’s anomalo- Venous blood was collected from the patients and available family scope. All color vision tests were performed under a ceiling-mounted members after informed consent and total genomic DNA was extracted lighting panel with four fluorescent tubes (36W/72 Biolux; Osram according to standard procedures. The samples were analyzed for GmbH, Munich, Germany), color temperature 6500 K. mutations in the following genes: CNGA3, CNGB3, GNAT2, KCNV2, Full-field electroretinography (ffERG) was performed in one eye in and PDE6C5,7–12 (SK, BW, unpublished data, 2009; details available on each of six patients after pupil dilation with cyclopentolate 1% and 30 request). Briefly, mutation screening was performed by PCR amplifi- minutes of dark adaptation according to the standard protocol of the cation from genomic DNA and sequencing of the coding exons and International Society for Clinical Electrophysiology of Vision flanking intron and UTR sequences. In addition, common alterations of (ISCEV).4,5 Briefly, the recordings where made with a Burian-Allen the red/green opsin OPN1MW/OPN1LW gene cluster typically found monopolar electrode after local anesthesia with oxybuprocaine 0.4%, a in blue cone monochromacy were excluded by means of PCR and reference ear clip electrode on the opposite earlobe, and a ground PCR/RFLP assays in male patients (data not shown). Segregation anal- electroencephalography silver cup electrode on the forehead. Stimu- ysis for the presence and independent inheritance of two mutant lations were produced with a xenon flash tube with adjustable inten- alleles was performed for the mutations c.886_896del11insT and sity mounted in a Ganzfeld globe (Nicolet Biomedical Instruments, c.1148delC in CNGB3 in both parents of patient D by PCR/RFLP Madison, WI). The flash intensity was 0.5 U in all recordings except analysis and direct sequencing. photopic red, green (both 1.25 U), and blue single flash (1.00 U). Flash duration was 20 to 30 ␮s. Flashes were attenuated with neutral-density filters. Data processing was performed on workstation (Viking IV; RESULTS Nicolet Biomedical Instruments). Reference values were obtained from a control group made up of 85 healthy adult volunteers. Ophthalmic Examination Multifocal electroretinography (mfERG; VERIS Science ver. 5.0; All patients had a history of congenital nystagmus and five Electro Diagnostic Imaging, Inc., San Mateo, CA) was recorded in five patients reported photophobia. Ophthalmic examination re- patients, with stimulus 103 hexagons at full contrast. A camera attach- vealed an unremarkable anterior segment in all six patients, ment was used for fixation, allowing eccentricity to be scaled. The disc drusen in patient B, and temporal peripapillary crescents stimulation pattern extended to 20° eccentricity. Data were processed and unilateral epiretinal fibrosis in patient A. Goldmann perim- with the manufacturer’s standard software. etry (IV/4e and I/4e) demonstrated no scotomata or constric- Macular thickness was measured by OCT in both eyes of five tions except that with the smaller target (I/4e) in patient D, in patients after pupil dilation (Stratus OCT model 3000, software ver. whom concentric constriction to 15° to 20° was seen, and in 4.0.1, and Spectral Domain Cirrus OCT; Carl Zeiss Meditec, Humphrey patient B, who was unable to see this target (Table 1). Five Division, Dublin, CA). A combination of fast and detailed protocols patients with a follow-up of eight or more years showed no were used to minimize the effect of nystagmus while obtaining high- sign of change in best corrected visual acuity, visual fields, or resolution tomograms for evaluation of retinal structure, and integral fundus morphology. All patients had subjectively normal or software was used to calculate retinal thickness and macular volume. near-normal color vision. In addition to reference data supplied by the manufacturer, we com- Color vision testing was normal for the Lanthony tritan pared the patients with 17 healthy adults and 1 child plus published album (n ϭ 5) and Farnsworth-Munsell’s D-15 standard test data from children.6 The OCT operator controlled centration on the (n ϭ 6; Table 2). Ishihara was read without errors by two fovea during the recording. Fixation errors deemed to be larger than patients, and four others had 1, 3, 5, and 8 errors, respectively 500 ␮m led to rejection of the scan. Morphologic evaluation was made (C, E, D, and B). Farnsworth-Munsell’s 100-hue test showed on the best-centered scans with the thinnest fovea and, if possible, some nonuniformity, with error scores ranging from a normal with a visible foveal reflex. score of 36 to a moderately abnormal score of 236 with no Fundus autofluorescence photography was performed in five pa- clear axis other than a probable tritan axis in patient D. tients after pupil dilation, with a confocal scanning laser ophthalmo- A recordable ffERG cone response, yet with amplitudes scope (Heidelberg Retina Angiograph; Heidelberg Engineering, Heidel- below those of the 5th percentile of healthy subjects and berg, Germany) set to align and average at least 16 frames. normal implicit times was found in patient A whose mfERG

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TABLE 2. Summary of Results of Color Vision Evaluation

Patient F-M 100 F-M D-15 LTA Ishihara 38-pl. Nagel Anomaloscope

A Error score 36, normal (normal No errors No errors No errors Normal (41–41/14) value for age: 100) B Error score 131, diffuse No errors No errors 8 errors — discrimination error (normal value for age: 100) C Error score 124, diffuse No errors No errors One error — discrimination error (normal value for age: 78) D Error score 236, diffuse Few neighbor No errors 5 errors — discrimination error, but transpositions probable tritan axis (normal value for age: 180) E Error score 67, normal (normal No errors One error 3 errors Normal with slightly value for age: 78) extended setting interval (38–43/14) F — Few neighbor — No errors — transpositions

—, Unavailable data.

responses were barely detectable (Fig. 1, Table 3). Cone re- Autofluorescence fundus images were normal in patients B, sponses of lower amplitude but still detectable were found in C, and E, with little enough nystagmus and photophobia to patients E and F. Cone responses were near the detection enable examination. threshold in the remaining three patients. Subnormal rod amplitudes were found in patients B, C, E, and F. Nearly absent Molecular Genetics mfERG cone responses were found in three patients, whereas mfERG failed in patient C and was unavailable in two patients. We analyzed the genes CNGA3, CNGB3, GNAT2, KCNV2, and Optical coherence tomography (n ϭ 4) demonstrated a PDE6C, which are all known to be involved in autosomal relative distribution of reflectivity among the retinal layers that recessive achromatopsia and cone dysfunction disorders, as was indistinguishable from that in healthy subjects, whereas well as common alterations in the OPN1MW/OPN1LW gene the overall level of reflectivity was lower than in healthy sub- cluster in male patients, typically found in blue cone mono- jects (Fig. 2). The thickness of the neurosensory retina was chromacy (Table 5). Patient D was a compound heterozygote normal in the fovea but 6.0% thinner than in healthy subjects with a deletion/insertion mutation c.886_896del11insT on the in the parafoveal region (P ϭ 0.0003, Table 4) and 5.8% thinner paternal allele and the 1-bp deletion c.1148delC on the mater- in the perifoveal region (P ϭ 0.0056). The foveal contour was nal allele in the CNGB3 gene (Table 5). The mutation mildly flattened (Fig. 2). One patient was unavailable for OCT c.886_896del11insT induces a frameshift downstream of examination, and in patient C, nystagmus prevented examina- Arg296 including a tail of eight novel amino acids followed by tion. a stop codon (p.Thr296TyrfsX9). The mutation c.1148delC

S N T I 0 2 4 6 8 10 12 14 16 18 20nV/deg^2

200 nV

0 80 ms

FIGURE 1. Multifocal electroretinogram with minimal cone responses from patient A. Patients B and E, with a degree of nystagmus that permitted examination, had comparable characteristics.

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TABLE 3. Amplitude and Implicit Time of Responses on ffERG

Patient (Normal Values) A B C D E F

Scotopic [Median amplitude (␮V) (5–95th percentile)/Implicit time (ms) (5–95th percentile)] Rod response b-wave [210 (124–302)/103 (81–123)] 162/112 74.5/86.5 89.9/101 204/102 100/132 80.7/142 Rod-cone response a-wave [205 (111–312)/23.0 (21.0–25.0)] 115/24.0 121/24.0 65.9/22.5 173/23.5 59.7/23.0 115/29.6 b-wave [427 (285–674)/48.0 (41.0–55.0)] 242/45.0 196/60.5 224/51.0 385/57.0 260/59.5 193.1/53.6 Oscillatory potential (OP2 amplitude) [77.0 (29.0–125.0)/25.2 (24.0–27.2)] 27.3/26.4 24.4/34.2 22.2/26.8 36.0/27.0 38.6/31.8 31.2/37.6 Flicker response [94 (53–145)/29.0 (27.0–34.0)] 42.0/27.0 000037.4/39.2 Photopic Single white flash response [136 (61–154)/29.0 (27.0–32.0)] 54.0/29.5 0 0 0 4.56/42.0 No data Flicker response [101 (61–154)/26.0 (25.0–30.0)] 51.7/27.0 0000Nodata Long-wave (red) single flash response [105.2 (64.5–172.9)/ 29.2 (27.0–32.0)] 51.2/29.8 000038.7/30.4 Middle-wave (green) single flash response [114.6 (62–181)/ 28.0 (26.0–32.2)] 52.7/28.8 000035.0/30.0 Short-wave (blue) single flash response [4.4 (0.9–8.9)/43.6 (39.4–51.0)] 00000 0

0, Absence of a recordable response.

results in a frameshift downstream of Thr383 and generates a c.1755GϾT p.Lys585Asn in PDE6C, respectively. No other premature stop codon after 12 altered amino acid residues mutation could be identiﬁed in the coding exons and ﬂanking (p.Thr383IlefsX13). Patients A and B were shown to carry intronic sequences of the respective genes in these two pa- single heterozygous missense mutations. Patient A carried the tients. The relevance of the mutations to the disease in these mutation c.1208GϾA p.Arg403Gln in CNGB3 and patient B patients remains unclear. The single mutation c.1208GϾA

FIGURE 2. Transfoveal optical coherence tomograms of the retina from a healthy subject (a, aged 11 years) and from two patients with OT (b, patient D; c, patient B). Pa- tients with OT with recordable tomograms were those who had the least nystagmus and these patients all demonstrated diffusely subnormal retinal reﬂectivity. NFL, nerve ﬁber layer; GCL, ganglion cell layer; IPL, inner plexiform layer; INL, inner nuclear layer; OPL, outer plexiform layer; ONL&HL, outer nuclear layer and Henle’s layer; ELM, external limiting membrane; IS, inner segment of photoreceptors; IS/OS, inner and outer segment junction of photoreceptors; OS, outer segments of photoreceptors; RPE, retinal pigment epithelium; OS/RPE, outer segment of photoreceptors junction with the RPE.

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TABLE 4. OCT Retinal Thickness of the Right Eye

Macular Volume Foveal Thickness Inner Peripheral Outer Peripheral (mm3) (␮m) Thickness (␮m)* Thickness (␮m)* Comments

Slight epiretinal Patient A 6.6 254 260.3 224.5 ﬁbrosis Patient B 6.7 179 244.5 239.3 Patient C 6.3 226 251.8 212.0 Age 12 Patient E 6.7 199 268.5 225.5 Mean value for patients 6.6 214.5 256.3 225.3 Normal value (SD)† 6.7 (1.13) 209.4 (55.15) 272.4 (31.25) 239.1 (45.39) P-value 0.0122 0.5532 0.0003 0.0056

* Mean retinal thickness by optical coherence tomography in relation to eccentricity (mean of temporal, superior, nasal, and inferior sectors in the patients’ right eyes). † Mean of 17 healthy adults.

p.Arg403Gln in CNGB3 is known to be associated with auto- other investigators1,14 that the condition may be underdiag- somal recessive achromatopsia, but also autosomal recessive nosed, as only ERG examinations in individuals with unex- cone dystrophy and autosomal recessive macular dystrophy.13 plained nystagmus and/or reduced visual acuity will reveal the The single mutation c.1755GϾT p.Lys585Asn in PDE6C is to condition. date unique to patient B. Mutations in PDE6C are a rare cause OT is characterized by the triad of low vision, severely of autosomal recessive achromatopsia (SK, BW, unpublished reduced ERG cone activity, and normal color vision. Concom- data, 2009). The genetic analysis of the OPN1MW/OPN1LW itant signs may include nystagmus and photophobia. Complete gene cluster on the X-chromosome in the male patients A, C, and incomplete achromatopsia and blue cone monochromacy and F confirmed its structural integrity, thereby excluding differ from OT by the severity of color vision impairment (e.g., genotypes typically associated with blue cone monochroma- Nagel anomaloscope identity over a wide range of green-red tism. The molecular genetic analysis in patients C, E, and F did mixtures with a steep linear sequence). However, it should be not result in the identification of any mutation or putative noted that the clinical characteristics of OT also may be pathogenic sequence variants in the analyzed genes CNGA3, present in two cone–rod dysfunctional entities, cone dystro- CNGB3, GNAT2, KCNV2, and PDE6C. phy with supernormal rod response15 and Åland eye disease, better known as incomplete congenital essential night blind- 16,17 DISCUSSION ness. Both conditions are characterized by distinct ERG characteristics that may escape attention: supernormal rod OT (OT) is a rare cone dysfunction with less than 30 cases responses, and a reduced b/a-wave proportion of the scotopic published so far. The six patients described herein and one mixed cone–rod response with cone flicker abnormalities. A described in a previous report5 constitute all cases found at our distinguishing feature of OT in relation to cone dystrophies and national low vision center within a limited number of birth cone–rod dystrophies is its appearance in early infancy and the cohorts in a population of approximately 6.0 million, leading to lack of progression. Our patients showed some clinical heter- a rough prevalence estimate of 1 in 500,000. We agree with ogeneity, especially regarding the results of color vision tests

TABLE 5. Genetic Analysis and Results

Patient CNGA3 CNGB3 GNAT2 KCNV2 PDE6C BCM

A Excluded c.1208GϾA p.Arg403Gln Excluded Excluded Excluded Normal OPN1MW/OPN1LW heterozygous gene cluster B Excluded Excluded Excluded Excluded c.1755GϾT NI (female) p.Lys585Asn heterozygous C Excluded Excluded Excluded Excluded Excluded Normal OPN1MW/OPN1LW gene cluster, but p.Cys203Arg in a distal gene* D NI 1. Allele: NI NI NI NI (female) c.886_896del11insT p.Arg296TyrfsX8/ 2. Allele: c.1148delC p.Thr383IlefsX12 E Excluded Excluded Excluded Excluded Excluded NI (female) F Excluded Excluded Excluded Excluded Excluded Normal OPN1MW/OPN1LW gene cluster

NI, not investigated. * The OPN1MW/OPN1LW gene cluster on the X-chromosome is typically composed of one proximal OPN1MW (M, middle-wave-sensitive opsin), and one or more distal OPN1LW (L, long-wave-sensitive opsin) genes. The analysis assesses the structural integrity of this gene cluster. In addition, a common functionally inactivating mutation c.607TϾC p.Cys203Arg is tested. Patient C was shown to carry an intact OPN1MW/OPN1LW cluster with a wild-type OPN1MW gene, as determined by OPN1MW-speciﬁc long-distance PCR and sequencing, and at least one OPN1LW gene. One of the OPN1LW genes was shown to carry the mutation c.607TϾC p.Cys203Arg. This genotype is not suggestive of a clinical diagnosis of blue cone monochromacy.

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and electroretinography. Mild color vision deficiency has been tween the two metal binding motifs, and affecting an amino reported with the OT phenotype1,5,18 as has variable degrees acid residue conserved between PDE6C proteins of various of residual cone function measured by ERG.1,14 Patients B, C, E, species. and F had subnormal rod specific responses. Similar changes Mutations in GNAT2, CNGA3, and CNGB3 are primarily have been reported in OT by others14 and also in achromatop- associated with autosomal recessive achromatopsia.7,9–11 Al- sia due to CNGA3 and CNGB3 mutations.2,13,19 though most of these patients were clinically diagnosed with Conceptually, OT has been regarded as an extreme variant complete achromatopsia, some patients showed residual cone of incomplete achromatopsia.20 However, until the molecular function, either electrophysically or psychophysically. There is basis and the pathophysiology of the condition is better under- evidence that certain mutations or combinations of mutations stood, a classification within a hierarchical system remains account for residual function.19,23 Indirectly, the involvement speculative. of these genes in OT supports the notion that this disorder is an Based on clinical observations, Van Lith1 hypothesized that extreme variant of incomplete achromatopsia. The large pro- the condition may be due to a reduced number of normal portion of cases with an essentially unknown molecular ge- functioning cones. This view was corroborated by the exper- netic background, however, indicates further genetic hetero- iments of Keunen et al.2 who, by foveal densitometry, demon- geneity, possibly involving different genetic mechanisms. strated a reduced density difference in the foveal cone In conclusion, this study confirmed the clinical findings photopigments in vivo and a normal time constant of photopig- overall comparable to that found by other investiga- ment regeneration. So far, no pathologic report exists, and it is tors.1,2,14,18,20,24 In addition, nearly absent mfERG responses, unknown whether the reduced cone responses are due to a grossly normal structure of the outer retina, and reduced peri- reduced number of cones or whether the majority of cones are foveal thickness on transfoveal OCT was found. Causative still present but morphologically and/or functionally silent. mutations have so far been identified in only two patients with The grossly normal structure of the outer retina on transfoveal OT.5 The negative and inconclusive outcome of the mutational OCT in this study does not suggest that a significant number of analyses in most of our patients gives indirect evidence of still cones are missing in OT. This observation is in contrast to unknown genes or genetic mechanisms being involved in OT. quantitative analysis of OCT characteristics in patients with achromatopsia and blue cone monochromacy. These analyses show absent IS/OS reflectivity in achromatopsia and a reduced Acknowledgments reflectivity with a reduced distance from the retinal pigment The authors thank the patients and family members for participating epithelium in blue cone monochromatism, indicating short- and Klaus Kallenbach and Cecilia Ro¨nnba¨ck for kindly providing the ened outer segments of the remaining photoreceptors.21 The OCT results from normal subjects. significance of our finding that the perifoveal retina is abnor- mally thin in OT is not known. We suggest that it may be caused by abnormal packing of the photoreceptors, ganglion References cells, or some other component of the retina during its devel- opment. Others have proposed a hypothetical entity of “pe- 1. Van Lith GHM. General cone dysfunction without achromatopsia. ripheral cone disease.”22 In: Pearlman JT, ed. 10th ISCERG Symposium. Documenta Oph- thalmologica Proceedings Series. Amsterdam: Kluwer Academic The molecular basis of OT is largely unknown. Compound Publishers; 1973;175–180. heterozygous mutations in GNAT2 were identified in a male 5 2. Keunen JEE, De Brabandere SRS, Liem ATA. Foveal densitometry patient with nystagmus and OT. One of the mutations, a deep and color mathing in oligocone trichromacy. In: Drum B, Moreland intron variation c.461ϩ24GϾA was shown to cause a splicing JD, Serra A, eds. 12th IRGCVD Symposium, Colour Vision Defi- defect that introduces a premature termination codon. Yet this ciencies XII. Amsterdam: Kluwer Academic Publishers; 1995:203– mutation was “leaky,” so that small amounts of correctly 210. spliced GNAT2 transcripts are formed and can eventually yield 3. Michaelides M, Hunt DM, Moore AT. The cone dysfunction syn- some functional protein. Moreover, compound heterozygous dromes. Br J Ophthalmol. 2004;88:291–297. CNGA3 mutations (p.Thr224Arg and p.Thr369Ser) were de- 4. Marmor MF, Fulton AB, Holder GE, Miyake Y, Brigell M, Bach M. scribed in two siblings with a mild form of incomplete achro- ISCEV Standard for full-field clinical electroretinography (2008 matopsia with moderate color vision deficiency.23 Functional update). Doc Ophthalmol. 2009;118(1):69–77. in vitro expression experiments of p.Thr224Arg mutant chan- 5. Rosenberg T, Baumann B, Kohl S, Zrenner E, Jorgensen AL, Wiss- inger B. Variant phenotypes of incomplete achromatopsia in two nels showed that it was a functional null mutation, whereas cousins with GNAT2 gene mutations. Invest Ophthalmol Vis Sci. p.Thr369Ser mutant channels were shown to form functional 2004;45:4256–4262. channels, albeit with some altered physiological properties, 6. Gupta G, Donahue JP, You T. Profile of the retina by optical that were partially compensated by coexpression with coherence tomography in the pediatric age group. Am J Ophthal- 23 CNGB3. mol. 2007;144:309–310. The compound heterozygous CNGB3 mutations in patient 7. Sundin OH, Yang JM, Li Y, et al. Genetic basis of total colourblind- D involve two frameshift mutations that are expected to give ness among the Pingelapese islanders. Nat Genet. 2000;25:289– rise to truncated proteins.11 Both mutations have been recur- 293. rently observed in patients with (complete) achromatopsia. It 8. Wissinger B, Gamer D, Jagle H, et al. CNGA3 mutations in hered- remains unclear why patient D expresses an OT phenotype, itary cone photoreceptor disorders. Am J Hum Genet. 2001;69: although she functionally performed the worst in the whole 722–737. patient series. Patients A and B were shown to carry single 9. Kohl S, Baumann B, Broghammer M, et al. Mutations in the CNGB3 heterozygous missense mutations. The relevance of the muta- gene encoding the beta-subunit of the cone photoreceptor cGMP- gated channel are responsible for achromatopsia (ACHM3) linked tions for the disease in these patients remains unresolved to chromosome 8q21. Hum Mol Genet. 2000;9:2107–2116. because of the absence of a second mutant allele. Patient A Ͼ 10. Kohl S, Baumann B, Rosenberg T, et al. Mutations in the cone carried the mutation c.1208G A p.Arg403Gln in CNGB3, photoreceptor G-protein alpha-subunit gene GNAT2 in patients which in the homozygous state causes autosomal recessive with achromatopsia. Am J Hum Genet. 2002;71:422–425. 14 progressive cone dystrophy. Patient B carried a single het- 11. Kohl S, Varsanyi B, Antunes GA, et al. CNGB3 mutations account erozygous mutation c.1755GϾT p.Lys585Asn in PDE6C, lo- for 50% of all cases with autosomal recessive achromatopsia. Eur cated in the catalytic domain of the PDE6C polypeptide be- J Hum Genet. 2005;13:302–308.

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